CN104431369B - Rumen-protected lysine feed - Google Patents
Rumen-protected lysine feed Download PDFInfo
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- CN104431369B CN104431369B CN201410778038.1A CN201410778038A CN104431369B CN 104431369 B CN104431369 B CN 104431369B CN 201410778038 A CN201410778038 A CN 201410778038A CN 104431369 B CN104431369 B CN 104431369B
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- feed addictive
- slug particle
- acquisition
- coating
- coated
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- 239000004472 Lysine Substances 0.000 title abstract description 25
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 title description 25
- PXEDJBXQKAGXNJ-QTNFYWBSSA-L disodium L-glutamate Chemical compound [Na+].[Na+].[O-]C(=O)[C@@H](N)CCC([O-])=O PXEDJBXQKAGXNJ-QTNFYWBSSA-L 0.000 claims abstract description 33
- 235000013923 monosodium glutamate Nutrition 0.000 claims abstract description 33
- 229940073490 sodium glutamate Drugs 0.000 claims abstract description 33
- 239000000945 filler Substances 0.000 claims abstract description 26
- 101710121765 Endo-1,4-beta-xylanase Proteins 0.000 claims abstract description 24
- 230000001070 adhesive effect Effects 0.000 claims abstract description 19
- 239000000853 adhesive Substances 0.000 claims abstract description 18
- 238000002360 preparation method Methods 0.000 claims abstract description 18
- 238000001035 drying Methods 0.000 claims abstract description 12
- 238000007873 sieving Methods 0.000 claims abstract description 10
- 239000002245 particle Substances 0.000 claims description 54
- 239000011248 coating agent Substances 0.000 claims description 53
- 238000000576 coating method Methods 0.000 claims description 53
- 230000000694 effects Effects 0.000 claims description 30
- 239000008187 granular material Substances 0.000 claims description 24
- 239000000203 mixture Substances 0.000 claims description 24
- 239000011122 softwood Substances 0.000 claims description 18
- 238000012216 screening Methods 0.000 claims description 11
- 229920001353 Dextrin Polymers 0.000 claims description 8
- 239000004375 Dextrin Substances 0.000 claims description 8
- 229920000168 Microcrystalline cellulose Polymers 0.000 claims description 8
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 8
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- KDXKERNSBIXSRK-YFKPBYRVSA-N L-lysine Chemical compound NCCCC[C@H](N)C(O)=O KDXKERNSBIXSRK-YFKPBYRVSA-N 0.000 abstract 3
- 239000011247 coating layer Substances 0.000 abstract 2
- 235000019766 L-Lysine Nutrition 0.000 abstract 1
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Landscapes
- Fodder In General (AREA)
Abstract
The invention provides a feed additive containing L-lysine. The feed additive consists of coated core particles and coating layers covering the coated core particles, wherein the coating layers are rumen-protected fat; and a preparation method of the coated core particles comprises the following steps: uniformly mixing lysine salt and a filler as well as optional sodium glutamate, adding an optional adhesive solution containing xylanase, sieving, shot-blasting, drying and sieving. In addition, the invention further provides a preparation method of the feed additive, wherein the preparation method comprises a step of spraying the rumen-protected fat on the coated core particles.
Description
Technical field
The invention belongs to amino acid field, specifically, feed addictive and its system the present invention relates to contain 1B
Preparation Method.
Background technology
1B is important amino acid starting material, can be used as flavouring, food, feed addictive, it is also possible to
As the effective or adjunct ingredient in health products, medicine, it is widely used in grocery trade, feed industry, pharmacy industry and other chemical works
In industry.Currently, the production of 1B mainly passes through the fermenting and producing of microorganism, can such as be produced using corynebacteria.
However, when lysine is as feed, because it belongs to small molecule nutriment, will soon be discharged into dynamic
In object, especially in huge uptake, the efficiency that animal absorbs is have impact on.The method for generally solving is to feed in multiple times on a small quantity
Food animal, but this will increase the workload of animal feeding.Therefore, the present invention is proposed and directly uses fermenting lysine nutrient solution system
The method of standby coated lysine product, can effectively slow down the speed that lysine discharges in animal body.
The preceding Chinese patent 201210440083 of the present inventor describes a kind of product of coated lysine, main to use
In feed.On this basis, present inventor has performed in-depth study, unexpectedly in many aspects, including process aspect,
Composition（Including trace mineral supplement）Aspect and enzyme transformation aspect etc., obtaining can improve what is be coated with and/or increase operation rate
The product of lysine.
Invention summary
The technical problem to be solved in the present invention is to provide new feed addictive containing 1B and preparation method thereof.
In addition, the present invention can also provide each branch invention obtained from feed addictive optimization, including the technique of optimization is being made
Application in the standby feed addictive containing 1B, a small amount of sodium glutamate is in the feed addictive containing 1B is prepared
Application, and/or, the application of the zytase of mutation in the feed addictive containing 1B is prepared etc..
Preferably, in one aspect of the invention, the invention provides the feed addictive containing 1B, it includes bag
By slug particle and the coated layer being wrapped on coating slug particle, wherein coated layer is rumen-undegraded fat, is coated with the preparation of slug particle
Method includes：
（1）Lysine salt and filler and optional sodium glutamate are well mixed, mixture is formed；
（2）To step（1）The optionally binder solution containing zytase is added in the mixture of acquisition, is well mixed,
Form softwood；
（3）By step（2）The softwood sieving of acquisition, forms coarse granule；
（4）By step（3）The coarse granule ball blast of acquisition, forms circular micropill；With
（5）Drying steps（4）The circular micropill of acquisition is simultaneously sieved, and forms coating slug particle.
Preferably wherein, step（1）In filler be microcrystalline cellulose and/or dextrin.
Preferably wherein, step（2）In adhesive be carboxymethylcellulose calcium.
Preferably wherein, step（2）In xylanase amino acid sequence such as SEQ ID NO：Shown in 2.
Preferably wherein, the weight ratio of lysine salt, filler, sodium glutamate and adhesive is 40 ~ 55：4~6：0~2：0.3~
0.8, preferably 42 ~ 50：4.5~5.5：0~1.5（Such as 0.5 ~ 1.5）：0.4 ~ 0.5, more preferably 43 ~ 48：4.8~5.2：0~1.2
（Such as 0.8 ~ 1.2）：0.45 ~ 0.55, most preferably 45 ~ 46：5：0~1：0.5.
Preferably wherein, the weight ratio of coating slug particle and coated layer is 40 ~ 60：40 ~ 60, preferably 46 ~ 56：43 ~ 53, more
Preferably 48.5 ~ 51.5：48.5 ~ 51.5, most preferably 1：1.
Preferably wherein, xylanase activity is 0 ~ 30IU, preferably 0 ~ 15IU in every gram of feed addictive（Such as 5 ~ 15IU）,
More preferably 0 ~ 10IU（Such as 8 ~ 10IU）, most preferably 0 ~ 9IU.
Preferably wherein, step（2）In sieve be 30 mesh sieves.
Preferably wherein, step（5）In screening be separate obtain can pass through 30 mesh sieves and by 40 mesh sieves retention particle.
On the other hand, the invention provides the preparation method of foregoing aspects of feed addictive, it is included rumen bypass
Fat is sprayed on the step on coating slug particle.
Detailed description of the invention
The technical problem to be solved in the present invention is to provide the new feed addictive containing 1B.In addition, of the invention
Can also provide the preparation method of the feed addictive and the technique of optimization prepare containing 1B feed addictive in
Application, a small amount of sodium glutamate prepare containing 1B feed addictive in application, and/or, the xylan of mutation
The invention of the branch such as enzyme and its application in the feed addictive containing 1B is prepared.
One, term is explained
Herein, term has the implication that those skilled in the art routinely understands.For the sake of clarity, herein
Following term is illustrated.
Herein, term is " optional " has its dictionary justice, i.e., the object being accordingly defined is chosen or does not select.Example
Such as, referred to containing containing optional sodium glutamate or do not contain sodium glutamate.And for example, optionally referred to containing zytase
Containing or do not contain zytase.
Herein, term " filler " and " adhesive " refer respectively to each serve as non-human animal's edible safety
Filling and the assistant agent of adhesive effect, it is preferable that " filler " and " adhesive " herein is food grade（That is people's edible safety
's）, it is referred to corresponding（People uses）The index of food.
Herein, term "comprising" or " including " be open statement, i.e., except list contain or comprise into
Outside part, can no longer contain other compositions or step, but be also not excluded for not influenceed containing other also the composition specifically listed or
Step plays the other compositions or step of function.Preferably, "comprising" herein or " including " be preferably " Consists of ".
Herein, as do not indicated plus especially, term " lysine " refers to 1B.
Two, the product containing 1B
Of the invention second（1）Aspect, the invention provides the feed addictive containing 1B, it includes being coated with core
Particle and the coated layer being wrapped on coating slug particle, wherein coated layer is rumen-undegraded fat, is coated with the preparation method of slug particle
Including：
（1）Lysine salt and filler and optional sodium glutamate are well mixed, mixture is formed；
（2）To step（1）The optionally binder solution containing zytase is added in the mixture of acquisition, is well mixed,
Form softwood；
（3）By step（2）The softwood sieving of acquisition, forms coarse granule；
（4）By step（3）The coarse granule ball blast of acquisition, forms circular micropill；With
（5）Drying steps（4）The circular micropill of acquisition is simultaneously sieved, and forms coating slug particle.
Preferably wherein, step（1）In filler be microcrystalline cellulose and/or dextrin.
Preferably wherein, step（2）In adhesive be carboxymethylcellulose calcium.
Preferably wherein, step（2）In xylanase amino acid sequence such as SEQ ID NO：Shown in 2.
Preferably wherein, the weight ratio of lysine salt, filler, sodium glutamate and adhesive is 40 ~ 55：4~6：0~2：0.3~
0.8, preferably 42 ~ 50：4.5~5.5：0~1.5（Such as 0.5 ~ 1.5）：0.4 ~ 0.5, more preferably 43 ~ 48：4.8~5.2：0~1.2
（Such as 0.8 ~ 1.2）：0.45 ~ 0.55, most preferably 45 ~ 46：5：0~1：0.5.
Preferably wherein, the weight ratio of coating slug particle and coated layer is 40 ~ 60：40 ~ 60, preferably 46 ~ 56：43 ~ 53, more
Preferably 48.5 ~ 51.5：48.5 ~ 51.5, most preferably 1：1.
Preferably wherein, xylanase activity is 0 ~ 30IU, preferably 0 ~ 15IU in every gram of feed addictive（Such as 5 ~ 15IU）,
More preferably 0 ~ 10IU（Such as 8 ~ 10IU）, most preferably 0 ~ 9IU.
Preferably wherein, step（2）In sieve be 30 mesh sieves.
Preferably wherein, step（5）In screening be separate obtain can pass through 30 mesh sieves and by 40 mesh sieves retention particle.
Of the invention second（2）Aspect, the invention provides of the invention second（1）The system of the feed addictive of aspect
Preparation Method, it includes the step being sprayed on rumen-undegraded fat on coating slug particle.
It is preferred that it includes：
（1）Lysine salt and filler and optional sodium glutamate are well mixed, mixture is formed；
（2）To step（1）The optionally binder solution containing zytase is added in the mixture of acquisition, is well mixed,
Form softwood；
（3）By step（2）The softwood sieving of acquisition, forms coarse granule；
（4）By step（3）The coarse granule ball blast of acquisition, forms circular micropill；
（5）Drying steps（4）The circular micropill of acquisition is simultaneously sieved, and forms coating slug particle；With
（6）Rumen-undegraded fat is sprayed on step（5）On the coating slug particle of acquisition.
Of the invention second（3）Aspect, the invention provides the product containing 1B, it include coating slug particle and
The coated layer on coating slug particle is wrapped in, wherein coated layer is rumen-undegraded fat, coating slug particle includes lysine salt, filling
Agent and adhesive and optional sodium glutamate and optional zytase.
Preferably wherein, product is feed addictive.
Preferably wherein, lysine salt is FE-5, lysine sulphate or their mixture, is preferably relied
Propylhomoserin sulfate.
Preferably wherein, filler is microcrystalline cellulose and/or dextrin.
Preferably wherein, adhesive is carboxymethylcellulose calcium.
Preferably wherein, xylanase amino acid sequence such as SEQ ID NO：Shown in 2.
Preferably wherein, the weight ratio of lysine salt, filler, sodium glutamate and adhesive is 40 ~ 55：4~6：0~2：0.3~
0.8, preferably 42 ~ 50：4.5~5.5：0~1.5（Such as 0.5 ~ 1.5）：0.4 ~ 0.5, more preferably 43 ~ 48：4.8~5.2：0~1.2
（Such as 0.8 ~ 1.2）：0.45 ~ 0.55, most preferably 45 ~ 46：5：0~1：0.5.
Preferably wherein, the weight ratio of coating slug particle and coated layer is 40 ~ 60：40 ~ 60, preferably 46 ~ 56：43 ~ 53, more
Preferably 48.5 ~ 51.5：48.5 ~ 51.5, most preferably 1：1.
Preferably wherein, often xylanase activity is 0 ~ 30IU, preferably 0 ~ 15IU in restraint product（Such as 5 ~ 15IU）, more preferably
It is 0 ~ 10IU（Such as 8 ~ 10IU）, most preferably 0 ~ 9IU.
Preferably wherein, coating slug particle is the particle that can pass through 30 mesh sieves and be retained by 40 mesh sieves.
Preferably wherein, fat is preferably palm fat and/or lecithin, the product that can be more preferably obtained by commercial sources
Product, such as hundred Canons, Pepsi U.S., preferably hundred Canon HTL316, Pepsi U.S. T300 etc..In specific embodiment of the invention
In, coating fat used is Pepsi U.S. T300.
Such as the instruction without opposite or contradiction, aspects above can preferably present aspect or other each side herein
Content.
Three, the technique for preparing the product containing 1B
The of the invention 3rd（1）Aspect, the invention provides the preparation method of the product containing 1B, it includes：
（1）Lysine salt and filler and optional sodium glutamate are well mixed, mixture is formed；
（2）To step（1）The optionally binder solution containing zytase is added in the mixture of acquisition, is well mixed,
Form softwood；
（3）By step（2）The softwood sieving of acquisition（Such as, 30 mesh sieve）, form coarse granule；
（4）By step（3）The coarse granule ball blast of acquisition, forms circular micropill；
（5）Drying steps（4）The circular micropill of acquisition is simultaneously sieved, and forms coating slug particle grain；With,
（6）Rumen-undegraded fat is sprayed on step（5）On the coating slug particle of acquisition.
It is preferred that the product containing 1B prepared by the preparation method is of the invention second（3）Product described in aspect.
It is preferred that the preparation method is made up of following steps：
（1）Lysine salt and filler and optional sodium glutamate are well mixed, mixture is formed；
（2）To step（1）The optionally binder solution containing zytase is added in the mixture of acquisition, is well mixed,
Form softwood；
（3）By step（2）The softwood sieving of acquisition（Such as, 30 mesh sieve）, form coarse granule；
（4）By step（3）The coarse granule ball blast of acquisition, forms circular micropill；
（5）Drying steps（4）The circular micropill of acquisition is simultaneously sieved, and forms coating slug particle；With,
（6）Rumen-undegraded fat is sprayed on step（5）On the coating slug particle of acquisition.
The inventors discovered that, the step of to coarse granule ball blast to form circular micropill and/or dry circle micropill and carry out
Screening, can effectively improve the anti-degradation capability of the coated product containing 1B of rumen-undegraded fat.So ensuing
Aspect, the invention provides the of the invention 3rd（1）Step in aspect（4）And/or step（5）Application.Throw preferably wherein
Ball is carried out in shot-blasting machine；And/or, screening is separated with the screen cloth of one or more special pore size distributions.
The of the invention 3rd（2）Aspect, the step of the invention provides to coarse granule ball blast to form circular micropill
Prepare the application in the method for the coated product containing 1B of rumen-undegraded fat.
The of the invention 3rd（3）Aspect, rumen bypass is being prepared the invention provides drying circle micropill and being sieved
Application in the method for the coated product containing 1B of fat.
The of the invention 3rd（4）Aspect, the step of the invention provides to coarse granule ball blast to form circular micropill
Improve the application in the anti-degradation capability of the coated product containing 1B of rumen-undegraded fat.
The of the invention 3rd（5）Aspect, is improving the invention provides dry circle micropill and the step of screening
Application in the anti-degradation capability of the coated product containing 1B of rumen-undegraded fat.
Preferably wherein, the step of to coarse granule ball blast to form circular micropill, is in（It is preferred that being next to）Formed coarse granule it
Afterwards.
Preferably wherein, dry circle micropill and be in the step of screening（It is preferred that being next to）Rumen-undegraded fat is sprayed on
Before on coating slug particle.
Preferably wherein, the coated product containing 1B of rumen-undegraded fat is of the invention second（3）Described in aspect
Product.
Preferably wherein, the method for preparing the coated product containing 1B of rumen-undegraded fat is the of the invention 3rd（1）
Method described in aspect.
Preferably wherein, screening is to separate the particle for obtaining and passing through 30 mesh sieves and be retained by 40 mesh sieves.
Such as the instruction without opposite or contradiction, aspects above can preferably present aspect or other each side herein
Content.
Four, application of the sodium glutamate in the product containing 1B
The inventors discovered that, a small amount of sodium glutamate is beneficial to the anti-degradation capability of the product containing 1B, and
Lysine is not influenceed to play the effect of feed.So, present invention also offers sodium glutamate in the product containing 1B
Using and correlation method.
The of the invention 4th（1）Aspect, the invention provides the preparation method of the product containing 1B, it includes：
（1）Lysine salt and filler and sodium glutamate are well mixed, mixture is formed；
（2）To step（1）The optionally binder solution containing zytase is added in the mixture of acquisition, is well mixed,
Form softwood；
（3）By step（2）The softwood sieving of acquisition（Such as, 30 mesh sieve）, form coarse granule；
（4）By step（3）The coarse granule ball blast of acquisition, forms circular micropill；
（5）Drying steps（4）The circular micropill of acquisition is simultaneously sieved, and forms coating slug particle；With,
（6）Rumen-undegraded fat is sprayed on step（5）On the coating slug particle of acquisition.
Preferably wherein, product is feed addictive.
Preferably wherein, step（1）In filler be microcrystalline cellulose and/or dextrin.
Preferably wherein, step（2）In adhesive be carboxymethylcellulose calcium.
Preferably wherein, step（2）In xylanase amino acid sequence such as SEQ ID NO：Shown in 2.
Preferably wherein, the weight ratio of lysine salt, filler, sodium glutamate and adhesive is 42 ~ 50：4.5~5.5：0.5~
1.5：0.4 ~ 0.5, more preferably 43 ~ 48：4.8~5.2：0.8~1.2：0.45 ~ 0.55, most preferably 45 ~ 46：5：1：0.5.
Preferably wherein, the weight ratio of coating slug particle and coated layer is 40 ~ 60：40 ~ 60, preferably 46 ~ 56：43 ~ 53, more
Preferably 48.5 ~ 51.5：48.5 ~ 51.5, most preferably 1：1.
Preferably wherein, xylanase activity is 0 ~ 30IU, preferably 0 ~ 15IU in every gram of feed addictive（Such as 5 ~ 15IU）,
More preferably 0 ~ 10IU（Such as 8 ~ 10IU）, most preferably 0 ~ 9IU.
Preferably wherein, xylanase activity is 0 ~ 30IU, preferably 0 ~ 15IU in every gram of feed addictive（Such as 5 ~ 15IU）,
More preferably 0 ~ 10IU（Such as 8 ~ 10IU）, most preferably 0 ~ 9IU.
Preferably wherein, step（5）In screening be separate obtain can pass through 30 mesh sieves and by 40 mesh sieves retention particle.
The of the invention 4th（2）Aspect, the invention provides the of the invention 4th（1）Prepared by the method described in aspect
Product.
The of the invention 4th（3）Aspect, the invention provides sodium glutamate to prepare rumen-undegraded fat coated containing L-
In the method for the product of lysine or in the anti-degradation capability for improving the coated product containing 1B of rumen-undegraded fat
Application.That is, the invention provides sodium glutamate in the method for preparing the coated product containing 1B of rumen-undegraded fat
Application, present invention also offers sodium glutamate improve the coated product containing 1B of rumen-undegraded fat anti-degraded energy
Application in power.
Preferably wherein, sodium glutamate is little, and the weight of such as sodium glutamate accounts for that rumen-undegraded fat is coated to be relied containing L-
The 0.5 ~ 1.5% of the product of propylhomoserin, preferably 0.8 ~ 1.2%, most preferably 1%.
Preferably wherein, the coated product containing 1B of rumen-undegraded fat is the of the invention 4th（2）Described in aspect
Product.
Preferably wherein, the method for preparing the coated product containing 1B of rumen-undegraded fat is the of the invention 4th（1）
Method described in aspect.
Such as the instruction without opposite or contradiction, aspects above can preferably present aspect or other each side herein
Content.
Five, the zytase of mutation and its application
The present inventor's surprisingly xylanase mutant, it is keeping the base of original high-temperature stability substantially
Enzymatic activity is added significantly on plinth, is particularly useful for being added into the feed addictive containing 1B.
The of the invention 5th（1）Aspect, the invention provides the zytase of mutation, it includes mutation Arg45Ser,
I.e. the 45th amino acids residue sports Ser by Arg.
It is preferred that the amino acid sequence of the zytase of the mutation
（1）Such as SEQ ID No:Shown in 2；Or
（2）It is to SEQ ID No:2 lack, add and/or replace one or several（Preferably more than 7, more preferably not
More than 5,3 are such as no more than）Amino acid residue and the reservation SEQ ID No that obtain:The sequence of 2 activity, and wherein include
Mutation Arg45Ser.
The of the invention 5th（2）Aspect, the invention provides coding the of the invention 5th（1）Xylan described in aspect
The gene of enzyme.
It is preferred that the nucleotide sequence of the gene such as SEQ ID No:Shown in 1.
The of the invention 5th（3）Aspect, the invention provides comprising the of the invention 5th（2）Gene described in aspect
Carrier.
The of the invention 5th（4）Aspect, the invention provides strain, it includes the of the invention 5th（2）Described in aspect
Gene, or with the of the invention 5th（3）Carrier described in aspect converts or transfects and obtain.
It is preferred that the strain is secreting, expressing bacterium, and e.g., yeast.
The of the invention 5th（5）Aspect, the invention provides fermenting and producing the of the invention 5th（1）Wood described in aspect
The method of dextranase, it includes cultivating the of the invention 5th under fermentation conditions（4）Strain described in aspect, divides from culture
Separate out the of the invention 5th（1）Zytase described in aspect.
The of the invention 5th（6）Aspect, the invention provides the of the invention 5th（1）Zytase described in aspect exists
Prepare the purposes in the product containing 1B.
Preferably wherein, the of the invention 5th in every gram of product containing 1B（1）Xylanase activity described in aspect
It is 5 ~ 15IU, more preferably 8 ~ 10IU, most preferably 9IU.
Preferably wherein, product is feed addictive.
Preferably wherein, the coated layer that the product containing 1B includes coating slug particle and is wrapped on coating slug particle,
Wherein coated layer is rumen-undegraded fat, and coating slug particle includes the of the invention 5th（1）Zytase, lysine described in aspect
Salt, filler and adhesive and optional sodium glutamate.
More preferably wherein, lysine salt is FE-5, lysine sulphate or their mixture, preferably
Lysine sulphate.
More preferably wherein, filler is microcrystalline cellulose and/or dextrin.
More preferably wherein, adhesive is carboxymethylcellulose calcium.
More preferably wherein, xylanase amino acid sequence such as SEQ ID NO：Shown in 2.
Preferably wherein, the weight ratio of lysine salt, filler, sodium glutamate and adhesive is 40 ~ 55：4~6：0~2：0.3~
0.8, preferably 42 ~ 50：4.5~5.5：0~1.5（Such as 0.5 ~ 1.5）：0.4 ~ 0.5, more preferably 43 ~ 48：4.8~5.2：0~1.2
（Such as 0.8 ~ 1.2）：0.45 ~ 0.55, most preferably 45 ~ 46：5：0~1：0.5.
Preferably wherein, the weight ratio of coating slug particle and coated layer is 40 ~ 60：40 ~ 60, preferably 46 ~ 56：43 ~ 53, more
Preferably 48.5 ~ 51.5：48.5 ~ 51.5, most preferably 1：1.
Preferably wherein, coating slug particle is the particle that can pass through 30 mesh sieves and be retained by 40 mesh sieves.
Preferably wherein, fat is preferably palm fat and/or lecithin, the product that can be more preferably obtained by commercial sources
Product, such as hundred Canons, Pepsi U.S., preferably hundred Canon HTL316, Pepsi U.S. T300 etc..In specific embodiment of the invention
In, coating fat used is Pepsi U.S. T300.
Preferably wherein, the method for preparation includes：
（1）Lysine salt and filler and optional sodium glutamate are well mixed, mixture is formed；
（2）To step（1）The binder solution containing zytase is added in the mixture of acquisition, is well mixed, formed
Softwood；
（3）By step（2）The softwood sieving of acquisition（Such as, 30 mesh sieve）, form coarse granule；
（4）By step（3）The coarse granule ball blast of acquisition, forms circular micropill；
（5）Drying steps（4）The circular micropill of acquisition is simultaneously sieved, and forms coating slug particle；With,
（6）Optionally, rumen-undegraded fat is sprayed on step（5）On the coating slug particle of acquisition.
Such as the instruction without opposite or contradiction, aspects above can preferably present aspect or other each side herein
Content.
The invention has the advantages that：By the improvement of many aspects, product safety reliability of the invention, relative to
Prior art, can more effectively reduce the rate of release of lysine in animal body, and anti-degradation capability is stronger, more improve the profit of feed
Promote growing for animal with rate.
In order to make it easy to understand, will be described in detail to the present invention by specific embodiment below.Needs are referred in particular to
Go out, the description that these descriptions are merely exemplary, and be not meant to limit the scope of the invention.According to the opinion of this specification
State, many changes of the invention, change will be apparent from for one of ordinary skill in the art.
In addition, the present invention refer to open source literature, these documents are their full text in order to more clearly describe the present invention
Content is included and referred to herein, just looks like that their full text is repeated and was expressly recited the same herein.
Specific embodiment
Present disclosure is further illustrated by the following examples.As do not specialized, technology used in embodiment
Conventional meanses and commercially available common instrument, reagent that means are well known to those skilled in the art, reference can be made to《Molecular Cloning: A Laboratory
Guide（3rd edition）》（Science Press）、《Microbiology Experiment（4th edition）》（Higher Education Publishing House）And corresponding instrument and
Manufacturers instruction of reagent etc. is referred to.
The preparation of the expression construct of the xylanase gene of the saltant type of embodiment 1
According to us in No. 201210185883 xylanase genes of record of preceding Chinese Patent No., the 45th will be thereon encoded
The Arg of position is mutated into Ser, will the CGC of nucleotide sequence 133-135 be mutated into TCC, the core of the zytase of saltant type
The SEQ ID No of nucleotide sequence such as sequence table:Shown in 1, the amino acid sequence such as SEQ ID No of coded zytase:2 institutes
Show.Then, according to《Molecular Cloning:A Laboratory guide》And the operating guidance of commercial reagents used builds the yeast of secreting, expressing,
In brief, the gene SEQ ID No of forward primer such as sequence table of the zytase will be encoded:Shown in 3（Introduce
EcoR I restriction enzyme sites）, reverse primer such as sequence table SEQ ID No:Shown in 4（Introduce Not I restriction enzyme sites）Expand
After increasing use EcoR I and Not I double digestions, and with the pPIC3.5 plasmids through the two endonuclease digestions（It is purchased from
Invitrogen companies）It is attached with T4 DNA ligases, is transformed into bacillus coli DH 5 alpha.Choose positive colony, extract
Go out plasmid therein, through sequence verification it is correct after, by positive plasmid with Sal I linearization for enzyme restriction after, by electrotransformation convert
Enter pichia yeast GS115 plants, positive yeast transformant is chosen on the flat board containing G418, return.
The fermentation preparation and determination of activity of the zytase of the saltant type of embodiment 2
The positive yeast transformant that embodiment 1 is obtained is inoculated into 50ml BMGY fluid nutrient mediums（100mM potassium phosphates delay
Fliud flushing（pH6.0）, wherein containing 1% yeast extract, 2% peptone, Yeast Nitrogen base 1.34%, biotin 4*10-5%, glucose
2%, glycerine 1%）In, cultivated to OD600 in 30 DEG C, 200rpm and reach 5.0.
Above-mentioned culture is transferred in 1L BMGY fluid nutrient mediums, is cultivated 24 hours in 30 DEG C, 200rpm, add 5ml
Methyl alcohol, then proceedes to be cultivated 8 days in 30 DEG C, 200rpm, and every 24 hours of period added 5ml methyl alcohol, ventilation control dissolved oxygen（DO）Greatly
In 20%, and it is 6.0 to control pH（Adjusted with ammoniacal liquor）.After the completion of culture, with 0.22 μm of membrane filtration and retain supernatant, on this
Clear liquid is detected through SDS-PAGE and is wherein rich in the corresponding protein of zytase, and thus fermentation obtains zytase supernatant.
Fresh supernatant is taken, is divided into three parts, portion refrigeration is stand-by（A）, potassium phosphate is used after a freeze-drying to solid
Buffer solution（pH6.0）Again dissolve（B）, it is a to use kaliumphosphate buffer after evaporation drying to solid in 80 DEG C of baking ovens
（pH6.0）Again dissolve（C）；Control uses phosphoric acid using No. 201210185883 xylanase genes of discovery of Chinese Patent No.
Potassium buffer solution（pH6.0）It is divided into three parts after dissolving, portion refrigeration is stand-by（CA）, another two parts of difference freeze-dryings（CB）With 80 DEG C of bakings
Case evaporation drying（CC）, kaliumphosphate buffer is then dissolved in again（pH6.0）.These xylanase activities are determined respectively, are converted into
The enzyme activity of original unit's volume supernatant or Unit Weight, calculates due to the enzyme activity loss late that high temperature dehydration is caused, as a result such as the institute of table 1
Show, show relative to it is original we have found that the zytase that is dehydrated of high temperature resistant, the zytase of saltant type of the invention, although
High temperature resistant water separation capability be slightly worse than it is original, but still remain more than 75% activity, especially surprisingly, enzyme activity
Property has and is obviously improved, even if by high temperature dehydration, the absolute value of its enzymatic activity is still above original zytase, or even
Higher than enzyme activity of original zytase when without high temperature dehydration.
The high temperature dehydration enzyme activity loss late of the zytase of the invention and commercially available of table 1
| Sample | Enzyme activity | Loss late | Sample | Enzyme activity | Loss late |
| A | 3.7*104IU/L | CA | 2.4*104IU/L | ||
| B | 3.5*104IU/L | 5.4% | CB | 2.2*104IU/L | 8.3% |
| C | 2.8*104IU/L | 24% | CC | 2.0*104IU/L | 17% |
The coating of the 1B of embodiment 3
According to the method that we record in preceding Chinese patent 201210440083, it is 63.5% to obtain 1B content（w/
w）The granular core containing 1B sulfate（Itself it is commercially pure 1B sulfate）, no longer with low-purity
1B outer layer covers, directly as the lysine raw material of feed of the invention.
The formula of the feed addictive of the invention of table 2
| Formula 1 | Formula 2 | Formula 3 | Formula 4 | |
| Lysine raw material | 46kg | 45kg | 46kg | 45kg |
| Sodium glutamate | / | 1kg | / | 1kg |
| Microcrystalline cellulose | 4kg | 4kg | 4kg | 4kg |
| Dextrin | 1kg | 1kg | 1kg | 1kg |
| Carboxymethylcellulose calcium（CMC） | 0.5 | 0.5 | 0.5 | 0.5 |
| Zytase | / | / | 9*105IU | 9*105IU |
| Rumen-undegraded fat powder | 48.5kg | 48.5kg | 48.5kg | 48.5kg |
As shown in table 2, detailed process is as follows for coating raw material used：
By the lysine raw material of formula ratio, microcrystalline cellulose and dextrin and（When formula is containing sodium glutamate）Paddy
Propylhomoserin sodium pours into trough type mixing machine, is well mixed, and obtains coating core mixture；
By the carboxymethylcellulose calcium of formula ratio 25L water（When formula does not contain zytase）Or according to implementation
Zytase supernatant prepared by the method for example 2（When formula is containing zytase）Immersion, is configured to 2% CMC solution,
Then directly pour into above-mentioned coating core mixture, be well mixed, form softwood；
Above-mentioned softwood is poured into swing comminutor, through 30 eye mesh screens（Aperture is 0.6mm）, it is made diameter and is about
The coarse granule of 0.6mm；
Above-mentioned coarse granule is poured into ball blast 3 minutes in the ball-type shot-blasting machine run with 2200rpm, circular micropill is formed；
Above-mentioned micropill is placed in the fluidized granulation seed-coating machine of 60 DEG C of EAT and is dried 45 minutes, then respectively through 30
Eye mesh screen（Aperture is 0.6mm）With 40 eye mesh screens（Aperture is 0.425mm）, retaining can be cut by 30 eye mesh screens and by 40 mu of screen clothes
The particle diameter for staying is the coating slug particle of 0.425-0.6mm；
The rumen-undegraded fat powder of formula ratio is placed in oil tank and is heated to 90 DEG C, until completely melted, in 80 DEG C of insulations simultaneously
Connect in the shower nozzle penetrating fluidized granulation seed-coating machine of fluidized granulation seed-coating machine, be sprayed on above-mentioned coating slug particle surface therein, its
Middle fluidized granulation seed-coating machine air-introduced machine rotating speed 3500rpm, 60 DEG C of EAT finally respectively obtains coating slug particle and rumen bypass
Fatty powder weight ratio about 1：1 above-mentioned four kinds of formulas distinguish corresponding coated 1B feed addictive.
In addition, being prepared substantially according to said process according to the formula of formula 1, different is to omit ball blast step, i.e.,
Directly coarse granule is placed in the fluidized granulation seed-coating machine of 60 DEG C of EAT and is dried, the 1B for obtaining control technique 1 is raised
Feed additives；
Formula according to formula 1 is prepared substantially according to said process, and different is two steps screening after omission is dried
Step, i.e., after being dried 45 minutes in the fluidized granulation seed-coating machine of 60 DEG C of EAT, do not sieve, directly by all of particle all
Proceed next step coating, obtain the 1B feed addictive of control technique 2.
The degraded test of the coated 1B feed addictive of embodiment 4
According to Zhao He etc.（Different proportion sheep's hay is Chinese with the research of ensilage dry matter degradability in cow rumen
Ox industry science, 34 (2):13-16）The internal Nylon Bag of description, it is each to what is prepared according to the method for embodiment 3 in adult dairy cattle
Kind coated feed addictive carries out degraded test, using wherein 1B as testing index.In the case of not coated,
The degradation rate of 1B is up to more than 95% after 2 hours（That is 1B conservation rate is less than 5%）, and coated feed addictive
Conservation rate is as shown in table 3 below：
The anti-degradation effect of the feed addictive of the invention of table 3
As shown in table 3, but if the method recorded in preceding Chinese patent 201210440083 relative to us, the present invention
Coating technique can further extend remaining time of the 1B in animal stomach, wherein whether with the addition of wood of the invention
Dextranase is little for encapsulating effect influence, although it was unexpectedly determined that ball blast and the technique of screening are very small, but adopt
Conservation rate about 10% can be improved after, more surprisingly, if wherein adding a small amount of sodium glutamate（If only added
Plus glutamic acid, effect will significantly be not so good as its sodium salt）, will cause that coated particle is not allowed more easily rupturable so that bad ammonia therein
Acid release is more smoothed, either in short term（As in 2 hours）Or it is long-term（Such as 24 or 48 hours）, all significantly improve lysine
Anti- degradation effect in cud.
Effectiveness of the coated 1B feed addictive of embodiment 5 to calf growth effect
Commission herding research institute of Ningxia academy of agricultural sciences is fed with March with all feeds additive prepared according to the method for embodiment 3
The calf in age 8 weeks, to be fed with without feed addictive of the invention（Only feed basal diet）With with the addition of commercially available non-packet
The equivalent lysine additives for forage of quilt is control.Result is as shown in table 4 below：
Growth effect of the feed addictive of the invention of table 4 to calf
| Feed | Initial average weight（kg） | Terminate average weight（kg） | Increased weight（kg） | Increased weight increase rate（%） |
| Basal diet | 125.7 | 165.3 | 39.6 | / |
| Basal diet+not coated lysine | 126.1 | 166.9 | 40.8 | 3.0 |
| Basal diet+formula 1 | 125.6 | 169.5 | 43.9 | 10.9 |
| Basal diet+formula 2 | 126.4 | 170.5 | 44.1 | 11.4 |
| Basal diet+formula 3 | 125.8 | 171.0 | 45.2 | 14.1 |
| Basal diet+formula 4 | 124.8 | 170.3 | 45.5 | 14.9 |
As shown in table 4, growth of the feeding 1B to calf has certain facilitation, but if directly feeds
Raise, its facilitation is very limited, only less than 5%, and the feed of equivalent lysine is added after the method for the present invention is coated with
Additive, can remarkably promote growth, improve more than 10%, after especially with the addition of zytase of the invention, more can be further
Promote growth about 4%.Due to being fed with frequency in experiment（Twice daily）It is higher, thus it is of the invention some formula longer time
Encapsulating conservation rate advantage embody not to be in the influence for growing calf very fully, but still old 0.5% and more than
Lifting, it is contemplated that it is actual raise in can embody more notable.In addition, during experiment, there is no adverse reaction to report, show the present invention
Formula be safe.
<110>Ningbo Eppen Biotech Co., Ltd.
<120>Rumen bypass lysine feed
<130>Electronic application
<160> 5
<170> PatentIn version 3.3
<210> 1
<211> 1428
<212> DNA
<213>Artificial sequence
<400> 1
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tttcagcgcg cgtccgatat ttatggcaaa gatggcctga gcccggaaga tcgcacccgc 180
gtgctggatc tgctgtttag cgatacccgc ggcgcgggcc tgaccattgt gcgcaacggc 240
attggcagca acagcaccat taaagatttt atgaacagca ttgaaccgtt tagcccgggc 300
agcccgagcg cgccgccgca ttatgtgtgg gatcgcaacg atagcggcca ggtgtggctg 360
agccatgaag cggcgagcta tggcgtgaac accttttatg cggatgcgtg gagcgcgccg 420
ggctatatga aaaccaacga tgatgatagc aacggcggct atctgtgcgg cgtgagcaac 480
accagctgcg cgagcggcga ttggaaacag gcgtatgcga actatctggt gcagtatatt 540
catttttatc agcaggtggg cattaaagtg acccatctgg gctttctgaa cgaaccgcag 600
gaagatgtga cctatgcgag catgctgagc gatggcaccc aggcggcggc ggattttatt 660
aaagtgctgg cgccgaccgt gaaagcggcg ggcctggatg tgaaactgac ctgctgcgat 720
ggcgtgggct gggaagaaca gcgcgcgatg ctgccgggcc tgcaggcggg cggcccggaa 780
catagcgcgg aaagctatct gagcgtgatt accgcgcatg gctataacag cccgccgacc 840
accccgctgg aaaccagcct gccggtgtgg atgaccgaat gggcggatct gaacgataac 900
tataccgcgg cgtggtatga aaacggcgcg ccgggcgaag gcctgacctg ggcgaaccgc 960
attcaggatg cgtttacccg cagcaacgtg agcgcgtttc tgcattggat tggcgcggaa 1020
aacggcacca gcaacagccc gctgattaac ctgaacggcg atagctatgt ggcgagcaaa 1080
cgcctgtggg cgtttggcca gtttagccgc tttgtgcgcc cgggcgcggt gcgcattgat 1140
gcggcgagca gcgatccgct ggtgaccgtg agcgcgtttc agaacaaaga aaacggcgtg 1200
gtggcgaccc aggtgattaa caacgcggat accgattatg aagtggaagt ggatctgacc 1260
ggccgcggcc atctggtgag cgtggtgcag ccgtatctga ccaacaacga acatgatctg 1320
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<211> 475
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Met Ile Ser Leu Ala Ala Ser Leu Pro Ile Pro Leu Val Ser Ala Leu
1 5 10 15
Pro His Ala His Ile Ser Val Asn Ala His Thr Arg Phe Gln Gln Val
20 25 30
Asp Gly Phe Gly Phe Ser Glu Ala Phe Gln Arg Ala Ser Asp Ile Tyr
35 40 45
Gly Lys Asp Gly Leu Ser Pro Glu Asp Arg Thr Arg Val Leu Asp Leu
50 55 60
Leu Phe Ser Asp Thr Arg Gly Ala Gly Leu Thr Ile Val Arg Asn Gly
65 70 75 80
Ile Gly Ser Asn Ser Thr Ile Lys Asp Phe Met Asn Ser Ile Glu Pro
85 90 95
Phe Ser Pro Gly Ser Pro Ser Ala Pro Pro His Tyr Val Trp Asp Arg
100 105 110
Asn Asp Ser Gly Gln Val Trp Leu Ser His Glu Ala Ala Ser Tyr Gly
115 120 125
Val Asn Thr Phe Tyr Ala Asp Ala Trp Ser Ala Pro Gly Tyr Met Lys
130 135 140
Thr Asn Asp Asp Asp Ser Asn Gly Gly Tyr Leu Cys Gly Val Ser Asn
145 150 155 160
Thr Ser Cys Ala Ser Gly Asp Trp Lys Gln Ala Tyr Ala Asn Tyr Leu
165 170 175
Val Gln Tyr Ile His Phe Tyr Gln Gln Val Gly Ile Lys Val Thr His
180 185 190
Leu Gly Phe Leu Asn Glu Pro Gln Glu Asp Val Thr Tyr Ala Ser Met
195 200 205
Leu Ser Asp Gly Thr Gln Ala Ala Ala Asp Phe Ile Lys Val Leu Ala
210 215 220
Pro Thr Val Lys Ala Ala Gly Leu Asp Val Lys Leu Thr Cys Cys Asp
225 230 235 240
Gly Val Gly Trp Glu Glu Gln Arg Ala Met Leu Pro Gly Leu Gln Ala
245 250 255
Gly Gly Pro Glu His Ser Ala Glu Ser Tyr Leu Ser Val Ile Thr Ala
260 265 270
His Gly Tyr Asn Ser Pro Pro Thr Thr Pro Leu Glu Thr Ser Leu Pro
275 280 285
Val Trp Met Thr Glu Trp Ala Asp Leu Asn Asp Asn Tyr Thr Ala Ala
290 295 300
Trp Tyr Glu Asn Gly Ala Pro Gly Glu Gly Leu Thr Trp Ala Asn Arg
305 310 315 320
Ile Gln Asp Ala Phe Thr Arg Ser Asn Val Ser Ala Phe Leu His Trp
325 330 335
Ile Gly Ala Glu Asn Gly Thr Ser Asn Ser Pro Leu Ile Asn Leu Asn
340 345 350
Gly Asp Ser Tyr Val Ala Ser Lys Arg Leu Trp Ala Phe Gly Gln Phe
355 360 365
Ser Arg Phe Val Arg Pro Gly Ala Val Arg Ile Asp Ala Ala Ser Ser
370 375 380
Asp Pro Leu Val Thr Val Ser Ala Phe Gln Asn Lys Glu Asn Gly Val
385 390 395 400
Val Ala Thr Gln Val Ile Asn Asn Ala Asp Thr Asp Tyr Glu Val Glu
405 410 415
Val Asp Leu Thr Gly Arg Gly His Leu Val Ser Val Val Gln Pro Tyr
420 425 430
Leu Thr Asn Asn Glu His Asp Leu Glu Ala Ser Ser Pro Ile Ile Thr
435 440 445
Leu Ala Leu Gly Ala Ala Thr Lys Phe Thr Ser Ile Val Pro Ala Arg
450 455 460
Ser Leu Val Ser Phe Val Ser Lys Thr Ala Leu
465 470 475
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<212> DNA
<213> Artificial
<220>
<223>Forward primer
<400> 3
cgaattcgat gattagcctg gc 22
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<212> DNA
<213> Artificial
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agcggccgca ttacagcgcg gtttt 25
Claims (16)
1. the feed addictive of 1B is contained, and it includes the coated layer for being coated with slug particle and being wrapped on coating slug particle, its
Middle coated layer is rumen-undegraded fat, and the preparation method for being coated with slug particle includes：
（1）Lysine salt and filler and sodium glutamate are well mixed, mixture is formed, wherein filler is microcrystalline cellulose
Element and/or dextrin；
（2）To step（1）The optionally binder solution containing zytase is added in the mixture of acquisition, is well mixed, formed
Softwood；
（3）By step（2）The softwood sieving of acquisition, forms coarse granule；
（4）By step（3）The coarse granule ball blast of acquisition, forms circular micropill；With
（5）Drying steps（4）The circular micropill of acquisition is simultaneously sieved, and forms coating slug particle,
Wherein, the weight ratio of lysine salt, filler, sodium glutamate and adhesive is 42 ~ 50：4.5~5.5：0.5~1.5：0.4~
0.5。
2. the feed addictive described in claim 1, wherein step（2）In adhesive be carboxymethylcellulose calcium.
3. the feed addictive described in claim 1, wherein step（2）In xylanase amino acid sequence such as SEQ ID NO：
Shown in 2.
4. the feed addictive described in claim 1, wherein, the weight ratio of lysine salt, filler, sodium glutamate and adhesive
It is 43 ~ 48：4.8~5.2：0.8~1.2：0.45~0.5.
5. the feed addictive described in claim 1, wherein the weight ratio of coating slug particle and coated layer is 40 ~ 60：40~60.
6. the feed addictive described in claim 1, wherein the weight ratio of coating slug particle and coated layer is 46 ~ 56：43~53.
7. the feed addictive described in claim 1, wherein the weight ratio of coating slug particle and coated layer is 48.5 ~ 51.5：
48.5~51.5。
8. the feed addictive described in claim 1, wherein the weight ratio of coating slug particle and coated layer is 1：1.
9. the feed addictive described in claim 1, wherein xylanase activity is 0 ~ 30IU in every gram of feed addictive.
10. the feed addictive described in claim 9, wherein xylanase activity is 0 ~ 15IU in every gram of feed addictive.
Feed addictive described in 11. claims 9, wherein xylanase activity is 5 ~ 15IU in every gram of feed addictive.
Feed addictive described in 12. claims 9, wherein xylanase activity is 0 ~ 10IU in every gram of feed addictive.
Feed addictive described in 13. claims 9, wherein xylanase activity is 8 ~ 10IU in every gram of feed addictive.
Feed addictive described in 14. claims 9, wherein xylanase activity is 0 ~ 9IU in every gram of feed addictive.
Feed addictive described in 15. claims 1, wherein step（3）In sieve be 30 mesh sieves.
Feed addictive described in 16. claims 1, wherein step（5）In screening be separate obtain can pass through 30 mesh sieves simultaneously
The particle retained by 40 mesh sieves.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
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